Supplemental Data
Files in this Data Supplement:
- Supplemental Figure 1 - QARIP analysis of type II membrane proteins detected in HEK293 cells. Protein topologies and peptide coverage are depicted for all type II membrane proteins listed in Tab. 1. Data analysis was performed and schemes shown were generated using QARIP (http://webclu.bio.wzw.tum.de/qarip). QARIP retrieves the topological information shown here either from Uniprot annotations of the respective protein entries or from Phobius predictions (http://phobius.sbc.su.se). Sequences covered by the peptides detected are indicated by black bars for tryptic peptides and pink bars for selected semitryptic peptides. Note that, with only one exception, peptides map exclusively to the extracellular/luminal domains.
- Supplemental Figure 2 - Immunoblot validation of the SPPL3 candidate substrate OGFOD3. Secreted (s) and cellular levels of OGFOD3 were monitored in TCA-precipitated supernatants (sup) and whole-cell lysates of HEK293 cells as detailed in Fig. 1.
- Supplemental Figure 3 - Immunoblot validation of the SPPL3 candidate substrates ASPH and TOR1AIP1. Secreted (s) and cellular levels of ASPH (A) and TOR1AIP1 (B) were monitored in TCA-precipitated supernatants (sup) and whole-cell lysates of HEK293 cells as detailed in Fig. 1. For both candidate substrates multiple bands (black arrowheads) were detected in lysates that were clearly specific as they were absent or reduced in cells transfected with siRNA targeting the respective candidate substrate. Secretion of ASPH- and TOR1AIP1-derived fragments was only apparent in supernatants of cells overexpressing SPPL3 (+Dox; grey arrowheads). For detection of TOR1AIP1 two distinct antibodies were used, one directed against an intracellular/nuclear epitope and one against the luminal domain of TOR1AIP (see sketch on the right part of panel B). Note, that secreted TOR1AIP1 was only detected with the pAb directed against the luminal domain.
- Supplemental Table 1 - Quantified glycoproteins of the SPPL3 overexpression secretome data set. Quantified glycoproteins of the SPPL3 overexpression secretome data set: Identified proteins in the protein groups output table of MaxQuant were filtered according the following criteria: glycoprotein according uniprot annotation, no contaminant according the contaminants file in Andromeda, at least 2 identified unique peptides. Furthermore the glycoprotein should be either detected in at least 4 out of 5 experiments in the wild type HEK cell condition or in at least 4 out of 5 experiments in the SPPL3 overexpression condition.
- Supplemental Table 2 - Unprocessed protein groups output table of the MaxQuant analysis of the SPPL3 overexpression secretome data set
- Supplemental Table 3 - Unprocessed peptides table of the MaxQuant Analysis of the SPPL3 overexpression secretome data set
- Supplemental Table 4 - Quantified glycoproteins of the SPPL3 knockout secretome data set. Quantified glycoproteins of the SPPL3 knockout secretome data set filtered according the following criteria: Identified proteins in the protein groups output table of MaxQuant were filtered according the following criteria: glycoprotein according uniprot annotation, no contaminant according the contaminants file in Andromeda, at least 2 unique peptides. Furthermore the glycoprotein should either be detected in at least 4 out of 5 experiments in the wild type MEF if the protein was strongly decreased in SPPL3 knockout condition or should be detected in at least 4 out of 5 experiments in SPPL3 knockout MEF if the protein was strongly increased in SPPL3 knockout condition.
- Supplemental Table 5 - Unprocessed protein groups output table of the MaxQuant analysis of the SPPL3 knockout secretome data set
- Supplemental Table 6 - Unprocessed peptides table of the MaxQuant Analysis of the SPPL3 knockout secretome data set
- Supplemental Table 7 - GO term Analysis of the significantly increased type-II membrane proteins in the SPPL3 overexpression secretome data set with Gorilla for function and process GO terms.